Plant Molecular Biology

, 68:173 | Cite as

Identification, cloning and characterization of a GDSL lipase secreted into the nectar of Jacaranda mimosifolia

  • Brian W. Kram
  • Elizabeth A. Bainbridge
  • M. Ann D. N. Perera
  • Clay Carter
Article

Abstract

The presence and function of several proteins secreted into floral nectars has been described in recent years. Here we report the presence of at least eight distinct proteins secreted into the floral nectar of the tropical tree Jacaranda mimosifolia (Bignoniaceae). Steps were initiated to identify and characterize these proteins in order to determine potential functions. The N-terminal sequence of the major Jacaranda nectar protein, JNP1, at 43 kDa contained similarity with members of the plant GDSL lipase/esterase gene family. Based upon this sequence, a full-length cDNA was isolated and predicted to encode a mature protein of 339 amino acids with a molecular mass of 37 kDa. Both raw nectar and heterologously expressed JNP1 displayed lipase/esterase activities. Interestingly, J. mimosifolia flowers produce an opaque, white colored nectar containing spherical, lipophilic particles approximately 5 μm in diameter and smaller. GS-MS analysis also identified the accumulation of free fatty acids within the nectar. It is proposed that JNP1 hydrolyzes Jacaranda nectar lipids with the concomitant release of free fatty acids. Potential functions of JNP1 in relation to pollinator attraction and prevention of microbial growth within nectar are briefly discussed.

Keywords

GDSL lipase Jacaranda mimosifolia Nectar Nectaries Nectary Pollination 

Abbreviations

JNP1

Jacaranda nectar protein 1

PNB

Para-nitrophenyl butyrate

SDS PAGE

Sodium dodecyl sulfate polyacrylamide gel electrophoresis

GC-MS

Gas chromatography-mass spectrometry

RT PCR

Reverse transcriptase polymerase chain reaction

References

  1. Akoh CC, Lee GC, Liaw YC, Huang TH, Shaw JF (2004) GDSL family of serine esterases/lipases. Prog Lipid Res 43:534–552PubMedCrossRefGoogle Scholar
  2. Altschul SF, Gish W, Miller W, Myers EW, Lipman DJ (1990) Basic local alignment search tool. J Mol Biol 215:403–410PubMedGoogle Scholar
  3. Baker H, Baker I (1973) Amino acids in nectar and their evolutionary significance. Nature 241:543–545CrossRefGoogle Scholar
  4. Baker H, Baker I (1975) Studies of nectar-constitution and pollinator–plant coevolution. In: Gilbert LE, Raven PH (eds) Coevolution of animals and plants. University of Texas Press, Austin, TX, pp 126–152Google Scholar
  5. Baker I, Baker HG (1976) Analyses of amino acids in flower nectars of hybrids and their parents, with phylogenetic implications. New Phytol 76:87–98CrossRefGoogle Scholar
  6. Baker H, Baker I (1983) A brief historical review of chemistry of floral nectar. In: Bentley BL (ed) The biology of nectaries. Columbia University Press, New York, pp 126–152Google Scholar
  7. Batra LR, Batra SWT, Bohart GE (1973) The mycoflora of domesticated and wild bees (Apoidea). Mycopathol Mycol Appl 49:13–44CrossRefGoogle Scholar
  8. Buchmann SL (1987) The ecology of oil flowers and their bees. Ann Rev Ecol Syst 18:343–369CrossRefGoogle Scholar
  9. Carter C, Thornburg RW (2000) Tobacco nectarin I. Purification and characterization as a germin-like, manganese superoxide dismutase implicated in the defense of floral reproductive tissues. J Biol Chem 275:36726–36733PubMedCrossRefGoogle Scholar
  10. Carter C, Thornburg RW (2004a) Is the nectar redox cycle a floral defense against microbial attack? Trends Plant Sci 9:320–324PubMedCrossRefGoogle Scholar
  11. Carter CJ, Thornburg RW (2004b) Tobacco nectarin III is a bifunctional enzyme with monodehydroascorbate reductase and carbonic anhydrase activities. Plant Mol Biol 54:415–425PubMedCrossRefGoogle Scholar
  12. Carter CJ, Thornburg RW (2004c) Tobacco nectarin V is a flavin-containing berberine bridge enzyme-like protein with glucose oxidase activity. Plant Physiol 134:460–469PubMedCrossRefGoogle Scholar
  13. Carter C, Graham RA, Thornburg RW (1999) Nectarin I is a novel, soluble germin-like protein expressed in the nectar of Nicotiana sp. Plant Mol Biol 41:207–216PubMedCrossRefGoogle Scholar
  14. Carter C, Healy R, O’Tool NM, Naqvi SM, Ren G, Park S, Beattie GA, Horner HT, Thornburg RW (2007) Tobacco nectaries express a novel NADPH oxidase implicated in the defense of floral reproductive tissues against microorganisms. Plant Physiol 143:389–399PubMedCrossRefGoogle Scholar
  15. CRC Handbook of Chemistry and Physics (2004) In: Lide, DR. (ed), CRC Press, Boca RatonGoogle Scholar
  16. Dalrymple BP, Cybinski DH, Layton I, McSweeney CS, Xue GP, Swadling YJ, Lowry JB (1997) Three Neocallimastix patriciarum esterases associated with the degradation of complex polysaccharides are members of a new family of hydrolases. Microbiology 143:2605–2614PubMedCrossRefGoogle Scholar
  17. Deans SG, Waterman PG (1993) Biological activity of volatile oils. In: Hay RKM, Waterman PG (eds) Volatile oil crops: their biology, biochemistry, and production. Longman Scientific and Technical, Essex, England, pp 97–111Google Scholar
  18. Deinzer M, Thomson P, Burgett D, Isaacson D (1977) Pyrrolizidine alkaloids: their occurrence in honey from tansy ragwort. Science 195:497–499PubMedCrossRefGoogle Scholar
  19. Dunford C, Cooper R, Molan P (2000) The use of honey in wound management. Nursing Standard 15:63–68PubMedGoogle Scholar
  20. Ecroyd CE, Franich RA, Kroese HW, Steward D (1995) Volatile constituents of Cactylanthus taylorii flower nectar in relation to flower pollination and browsing by animals. Phytochemistry 40:1387–1389CrossRefGoogle Scholar
  21. Ferreres F, Andrade P, Gil MI, Tomas Barberan FA (1996) Floral nectar phenolics as biochemical markers for the botanical origin of heather honey. Z Lebensm Unters Forsch 202:40–44CrossRefGoogle Scholar
  22. Fiehn O, Kopka J, Trethewey RN, Willmitzer L (2000) Identification of uncommon plant metabolites based on calculation of elemental compositions using gas chromatography and quadrupole mass spectrometry. Anal Chem 72:3573–3580PubMedCrossRefGoogle Scholar
  23. Galetto L (1994) Nectary structure and nectar characteristics in some Bignoniaceae. Plant System Evol 196:1615–6110Google Scholar
  24. Griebel C, Hess G (1990) The vitamin C content of flower nectar of certain Labiatae. Z Unters Lebensm 79:168–171CrossRefGoogle Scholar
  25. Heinrich G (1989) Analysis of cations in nectars by means of a laser microprobe mass analyser (LAMMA). Beitr Biol Pflanz 64:293–308Google Scholar
  26. Hildebrand EM (1937) The blossomblight phase of fire blight, and methods of control. Cornell Univ Agric Exp Station Mem 207:1–40Google Scholar
  27. Hong JK, Choi HW, Hwang IS, Kim DS, Kim NH, Choi DS, Kim YJ, Hwang BK (2008) Function of a novel GDSL-type pepper lipase gene, CaGLIP1, in disease susceptibility and abiotic stress tolerance. Planta 227:539–558PubMedCrossRefGoogle Scholar
  28. Hopkins CY, Jevans AW, Bock R (1969) Occurrence of octadecatrans-2,cis-9,cis-12 trienoic acid in pollen attractive to the honey bee. Can J Biochem 47:433–436PubMedCrossRefGoogle Scholar
  29. Jaikaran AS, Kennedy TD, Dratewka-Kos E, Lane BG (1990) Covalently bonded and adventitious glycans in germin. J Biol Chem 265(21):12503–12512PubMedGoogle Scholar
  30. Kevan PG, Eisikowitch D, Fowle S, Thomas K (1988) Yeast-contaminated nectar and its effects on bee foraging. J Apicult Res 27:26–29Google Scholar
  31. Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685PubMedCrossRefGoogle Scholar
  32. Lepage M, Boch R (1968) Pollen lipids attractive to honeybees. Lipids 3:530–534PubMedCrossRefGoogle Scholar
  33. Li J, Derewenda U, Dautzer Z, Smith S, Derewenda ZS (2000) Crystal structure of the Escherichia coli thioesterase II, a homolog of the human Nef binding enzyme. Nat Struct Biol 7:555–559PubMedCrossRefGoogle Scholar
  34. Liu M, Gonzalez JE, Willis LB, Walker GC (1998) A novel screening method for isolating exopolysaccharide-deficient mutants. Appl Environ Microbiol 64:4600–4602PubMedGoogle Scholar
  35. Molgaard A, Kauppinen S, Larsen S (2000) Rhamnogalacturonan acetylesterase elucidates the structure and function of a new family of hydrolases. Structure 8:373–383PubMedCrossRefGoogle Scholar
  36. Naqvi SM, Harper A, Carter C, Ren G, Guirgis A, York WS, Thornburg RW (2005) Nectarin IV, a potent endoglucanase inhibitor secreted into the nectar of ornamental tobacco plants. Isolation, cloning, and characterization. Plant Physiol 139:1389–1400PubMedCrossRefGoogle Scholar
  37. Oh IS, Park AR, Bae MS, Kwon SJ, Kim YS, Lee JE, Kang NY, Lee S, Cheong H, Park OK (2005) Secretome analysis reveals an Arabidopsis lipase involved in defense against Alternaria brassicicola. Plant Cell 17:2832–2847PubMedCrossRefGoogle Scholar
  38. Peumans WJ, Smeets K, Van Nerum K, Van Leuven F, Van Damme EJ (1997) Lectin and alliinase are the predominant proteins in nectar from leek (Allium porrum L.) flowers. Planta 201:298–302PubMedCrossRefGoogle Scholar
  39. Pleasants JM, Chaplin SJ (1983) Nectar production rates of Asclepias quadrifolia: causes and consequences of individual variation. Oecologia 59:232–238CrossRefGoogle Scholar
  40. Purdy RE, Kolattukudy PE (1973) Depolymerization of a hydroxy fatty acid biopolymer, cutin, by an extracellular enzyme from Fusarium solani f. pisi: isolation and some properties of the enzyme. Arch Biochem Biophys 159:61–69PubMedCrossRefGoogle Scholar
  41. Roshchina VV, Roshchina VD (1993) The excretory function of higher plants. Springer-Verlag, New YorkGoogle Scholar
  42. Sambrook J, Russel DW (2001) Molecular cloning. Cold Spring Harbor Press, Cold Spring Harbor, New YorkGoogle Scholar
  43. Sommer P, Bormann C, Gotz F (1997) Genetic and biochemical characterization of a new extracellular lipase from Streptomyces cinnamomeus. Appl Environ Microbiol 63:3553–3560PubMedGoogle Scholar
  44. Taipalensuu J, Falk A, Rask L (1996) A wound- and methyl jasmonate-inducible transcript coding for a myrosinase-associated protein with similarities to an early nodulin. Plant Physiol 110:483–491PubMedCrossRefGoogle Scholar
  45. Tholl D, Chen F, Petri J, Gershenzon J, Pichersky E (2005) Two sesquiterpene synthases are responsible for the complex mixture of sesquiterpenes emitted from Arabidopsis flowers. Plant J 42:757–771PubMedCrossRefGoogle Scholar
  46. Thornburg RW, Carter C, Powell A, Mittler R, Rizhsky L, Horner HT (2003) A major function of the tobacco floral nectary is defense against microbial attack. Plant Syst Evol 238:211–218Google Scholar
  47. Upton C, Buckley JT (1995) A new family of lipolytic enzymes? Trends Biochem Sci 20:178–179PubMedCrossRefGoogle Scholar
  48. Vitale A, Chrispeels MJ (1992) Sorting of proteins to the vacuoles of plant cells. Bioessays 14:151–160PubMedCrossRefGoogle Scholar
  49. Vogel S (1969) Flowers offering fatty oil instead of nectar. In: Abstracts XIth International Botany Congress, Seattle, WAGoogle Scholar
  50. Zhang Z, Ober JA, Kliebenstein DJ (2006) The gene controlling the quantitative trait locus EPITHIOSPECIFIER MODIFIER1 alters glucosinolate hydrolysis and insect resistance in Arabidopsis. Plant Cell 18:1524–1536PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2008

Authors and Affiliations

  • Brian W. Kram
    • 1
  • Elizabeth A. Bainbridge
    • 2
  • M. Ann D. N. Perera
    • 2
  • Clay Carter
    • 1
  1. 1.Department of BiologyUniversity of Minnesota DuluthDuluthUSA
  2. 2.W.M. Keck Metabolomics Research LaboratoryIowa State UniversityAmesUSA

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